Organisms eliminate unwanted cells by a process variously known as regulated cell death, programmed cell death or apoptosis. Such cell death occurs as a normal aspect of animal development, as well as in tissue homeostasis and aging (Glucksmann, A., Biol. Rev. Cambridge Philos. Soc. 26:59-86 (1951); Glucksmann, A., Archives de Biologie 76:419-437 (1965); Ellis, et al., Dev. 112:591-603 (1991); Vaux, et al., Cell 76:777-779 (1994)). Apoptosis regulates cell number, facilitates morphogenesis, removes harmful or otherwise abnormal cells and eliminates cells that have already performed their function. Additionally, apoptosis occurs in response to various physiological stresses, such as hypoxia or ischemia (PCT published application WO96/20721).
There are a number of morphological changes shared by cells experiencing regulated cell death, including plasma and nuclear membrane blebbing, cell shrinkage (condensation of nucleoplasm and cytoplasm), organelle relocalization and compaction, chromatin condensation and production of apoptotic bodies (membrane enclosed particles containing intracellular material) (Orrenius, S., J. Internal Medicine 237:529-536 (1995)).
Apoptosis is achieved through an endogenous mechanism of cellular suicide (Wyllie, A. H., in Cell Death in Biology and Pathology, Bowen and Lockshin, eds., Chapman and Hall (1981), pp. 9-34). A cell activates its internally encoded suicide program as a result of either internal or external signals. The suicide program is executed through the activation of a carefully regulated genetic program (Wyllie, et al., Int. Rev. Cyt. 68:251 (1980); Ellis, et al., Ann. Rev. Cell Bio. 7:663 (1991)). Apoptotic cells and bodies are usually recognized and cleared by neighboring cells or macrophages before lysis. Because of this clearance mechanism, inflammation is not induced despite the clearance of great numbers of cells (Orrenius, S., J. Internal Medicine 237:529-536 (1995)).
It has been found that a group of proteases are a key element in apoptosis (see, e.g., Thornberry, Chemistry and Biology 5:R97-R103 (1998); Thornberry, British Med. Bull. 53:478-490 (1996)). Genetic studies in the nematode Caenorhabditis elegans revealed that apoptotic cell death involves at least 14 genes, 2 of which are the pro-apoptotic (death-promoting) ced (for cell death abnormal) genes, ced-3 and ced-4. CED-3 is homologous to interleukin 1 beta-converting enzyme, a cysteine protease, which is now called caspase-1. When these data were ultimately applied to mammals, and upon further extensive investigation, it was found that the mammalian apoptosis system appears to involve a cascade of caspases, or a system that behaves like a cascade of caspases. At present, the caspase family of cysteine proteases comprises 14 different members, and more may be discovered in the future. All known caspases are synthesized as zymogens that require cleavage at an aspartyl residue prior to forming the active enzyme. Thus, caspases are capable of activating other caspases, in the manner of an amplifying cascade.
Apoptosis and caspases are thought to be crucial in the development of cancer (Apoptosis and Cancer Chemotherapy, Hickman and Dive, eds., Humana Press (1999)). There is mounting evidence that cancer cells, while containing caspases, lack parts of the molecular machinery that activates the caspase cascade. This makes the cancer cells lose their capacity to undergo cellular suicide and the cells become cancerous. In the case of the apoptosis process, Control points are known to exist that represent points for intervention leading to activation. These control points include the CED-9-BCL-like and CED-3-ICE-like gene family products, which are intrinsic proteins regulating the decision of a cell to survive or die and executing part of the cell death process itself, respectively (see, Schmitt, et al., Biochem. Cell. Biol. 75:301-314 (1997)). BCL-like proteins include BCL-xL and BAX-alpha, which appear to function upstream of caspase activation. BCL-xL appears to prevent activation of the apoptotic protease cascade, whereas BAX-alpha accelerates activation of the apoptotic protease cascade.
It has been shown that chemotherapeutic (anti-cancer) drugs can trigger cancer cells to undergo suicide by activating the dormant caspase cascade. This may be a crucial aspect of the mode of action of most, if not all, known anticancer drugs (Los, et al., Blood 90:3118-3129 (1997); Friesen, et al., Nat. Med. 2:574 (1996)). The mechanism of action of current antineoplastic drugs frequently involves an attack at specific phases of the cell cycle. In brief, the cell cycle refers to the stages through which cells normally progress during their lifetime. Normally, cells exist in a resting phase termed Go. During multiplication, cells progress to a stage in which DNA synthesis occurs, termed S. Later, cell division, or mitosis occurs, in a phase called M. Antineoplastic drugs, such as cytosine arabinoside, hydroxyurea, 6-mercaptopurine, and methotrexate are S phase specific, whereas antineoplastic drugs, such as vincristine, vinblastine, and paclitaxel are M phase specific. M phase specific antineoplastic drugs, such as vinblastine and paclitaxel, are known to affect tubulin polymerization. The ability of cells to appropriately polymerize and depolymerize tubulin is thought to be an important activity for M phase cell division.
Many slow growing tumors, e.g. colon cancers, exist primarily in the Go phase, whereas rapidly proliferating normal tissues, for example bone marrow, exist primarily in the S or M phase. Thus, a drug like 6-mercaptopurine can cause bone marrow toxicity while remaining ineffective for a slow growing tumor. Further aspects of the chemotherapy of neoplastic diseases are known to those skilled in the art (see, e.g., Hardman, et al., eds., Goodman and Gilman's The Pharmacological Basis of Therapeutics, Ninth Edition, McGraw-Hill, New York (1996), pp. 1225-1287). Thus, it is clear that the possibility exists for the activation of the caspase cascade, although the exact mechanisms for doing so are not clear at this point. It is equally clear that insufficient activity of the caspase cascade and consequent apoptotic events are implicated in various types of cancer. The development of caspase cascade activators and inducers of apoptosis is a highly desirable goal in the development of therapeutically effective antineoplastic agents. Moreover, since autoimmune disease and certain degenerative diseases also involve the proliferation of abnormal cells, therapeutic treatment for these diseases could also involve the enhancement of the apoptotic process through the administration of appropriate caspase cascade activators and inducers of apoptosis.
EP520722 discloses derivatives of 4-anilino-quinazolines as inhibitors of the EGFR tyrosine kinase with antitumor activity:
wherein, for example, Ra is hydrogen, trifluoromethyl, or nitro, n is 1; and Rb is halogen, trifluoromethyl or nitro.
EP602851 discloses quinazolines as inhibitors of the EGFR tyrosine kinase:
wherein, for example Ra is hydroxy, amino, ureido, or trifluoromethoxy, m is 1, 2 or 3; Q is a 9 or 10-membered bicyclic heterocyclic moiety.
EP635498 discloses 4-anilino-quinazolines as inhibitors of the EGFR tyrosine kinase:
wherein, for example R1 includes hydroxy, amino or C1-4 alkoxy, R2 is hydrogen, hydroxy, or halogen, R3 is halogen, n is 1, 2 or 3.
EP635507 discloses tricyclic derivatives as inhibitors of the EGFR tyrosine kinase:
wherein, R1 and R2 together form an optionally substituted 5 or 6 membered ring containing at least one heteroatom; R3 includes hydrogen, hydroxy, or halogen, m is 1, 2 or 3.
WO9609294 discloses substituted heteroaromatic compounds as inhibitors of protein tyrosine kinase:
wherein, for example X is N or CH; Y is O, S, or NRa wherein Ra is H or C1-8 alkyl; R1, R2, R3 and R3′ includes amino, hydrogen, hydroxy, or halogen; R4 includes amino, hydrogen, hydroxy, or halogen; n is 1, 2 or 3; R5 is selected from the group comprising hydrogen, halogen, trifluoromethyl, C1-4 alkyl and C1-4 alkoxy; R6 is a group ZR7 wherein Z includes O, S or NH and R7 is an optionally substituted C3-6 cycloalkyl, or an optionally substituted 5,6,7,8,9,10-membered carbocyclic or heterocyclic moiety.
WO9713771 discloses substituted heteroaromatic compounds as inhibitors of protein tyrosine kinase:
wherein, for example X is N or CH; U represents a fused 5,6,7-membered heterocyclic ring; Y is O, S, or NRa wherein Ra is H or C1-8 alkyl; R1 included 5,6-membered heterocyclic ring, or amino, hydrogen, hydroxy, or halogen; n is 0, 1, 2 or 3. R2 is selected from the group comprising hydrogen, halogen, trifluoromethyl, C1-4 alkyl and C1-4 alkoxy; R3 is a group ZR4 wherein Z includes O, S or NH and R4 is an optionally substituted C3-6 cycloalkyl, or an optionally substituted 5,6,7,8,9,10-membered carbocyclic or heterocyclic moiety. R5 includes hydrogen, hydroxy, or halogen; n is 1, 2 or 3.
WO9802438 discloses bicyclic heteroaromatic compounds as inhibitors of protein tyrosine kinase:
wherein, for example X is N or CH; Y is O, S, or NRa wherein Ra is H or C1-8 alkyl; R″ represents a phenyl group or a 5- or 6-membered heterocyclic ring, or amino, hydrogen, hydroxy, or halogen; n is 0 or 1. R1 includes amino, hydrogen, hydroxy, or halogen; p is 0 to 3. R2 is selected from the group comprising hydrogen, halogen, trifluoromethyl, C1-4 alkyl and C1-4 alkoxy; U represents a 5 to 10-membered mono or bicyclic ring system; A represents a fused 5, 6, or 7-membered heterocyclic ring.
Myers et al. (Bioorg. Med. Chem. Lett. 7:421-424 (1997)) reported 4-(N-methyl-N-phenyl)amino-6,7-dimethoxyquinazoline as inhibitor of CSF-1R tyrosine kinase. It was reported that substitutions on the phenyl ring resulted in reduced activity. Replacement of the 6,7-dimethoxy groups by hydrogen resulted in more than 40-fold reduction in potency. Substitution in the 2-position of quinazoline by a Cl or methoxy group resulted in inactive compounds (IC50>50 μM).

Rewcastle et al. (J. Med. Chem. 38:3482-3487 (1995)) reported 4-(phenylamino)-quinazolines as inhibitors of tyrosine kinase of Epidermal Growth Factor Receptor. It was reported that N-methylation of the amino group (R1═Me, R2═R3═R4═H) completely abolished activity (IC50>100,000 nM). The 6,7-dimethoxy compound (R1═H, R2═R3═OMe, R4═Br, IC50=0.029 nM) was almost 1000-fold more potent than the corresponding non-substituted analog (R1═H, R2═R3═H, R4═Br, IC50=27 nM).

Bridges et al. (J. Med. Chem. 39:267-276 (1996)) reported analogs of 4-(3-bromoanilino)-6,7-dimethoxyquinazoline as inhibitors of tyrosine kinase of Epidermal Growth Factor Receptor. It was reported that introduction of a methyl group to the 2-position (R1═Me, R2=3′-Br, R3═H) resulted in at least 400,000-fold loss of potency (IC50>10,000 nM) vs the hydrogen analog. Introduction of an amino group to the 2-position (R1═NH2, R2=3′-Br, R3═H) also resulted in over 18,000-fold loss of potency (IC50>10,000 nM). Methylation of the anilino nitrogen (R3═Me) led to 6,000-fold drop in activity. The 4′-Br analog (IC50=0.96 nM) was almost 40-fold less active than the 3′-Br analog (IC50=0.025 nM), and the 2′-Br analog (IC50=128 nM) was at least 5,000-fold less active than the 3′-Br analog.
